| The pollution of water resources seriously threatens human health and sustainable development of society.For example,breeding wastewater is difficult to treat because it has the characteristics of large discharge,high organic pollutants content,and many kinds of pathogenic bacteria.In addition,existence of heterocyclic compounds in industrial wastewater causes pollution to groundwater,river sediments,seawater,and soil.Different types of wastewaters pose a threat to the environment and human health,so the effective treatment of wastewater remains a current concern.Therefore,establishing an effective wastewater treatment method is still a great challenge for the development of society.Microorganisms play an important role in the biodegradation of organic pollutants,chemical element recycling,and energy regeneration.Microbial degradation technology has become an active field in the treatment of organic pollutants due to its advantages of economy,high efficiency.convenient operation,and no secondary pollution.Microbial fuel cell(MFC)is a device that combines traditional pollutant degradation with electrochemical technology,and can use microorganisms to directly convert chemical energy in organic matter into electrical energy.Extracellular electron transfer(EET)of electroactive microorganisms is the basic working principle of MFC to recover resources from pollutants,which can promote the degradation of organic pollutants,the recovery of various elements,and the immobilization of heavy metals,and radioactive wastes.Therefore.MFC as a platform by utilizing microorganisms with EET capability can simultaneously achieve pollutant degradation and energy recovery.It is of great significance to alleviate energy crisis and solve environmental pollution.In this research,MFC was firstly used to treat breeding wastewater to evaluate the feasibility of it as a treatment device for wastewater.The obtained microbial community composition changes can illustrate the role of electroactive microorganisms in the wastewater treatment process.Because there are many kinds of microorganisms in wastewater,it is a difficulty to study the mechanism of EET in MFC.Therefore.Sphingobirm yanoikuyae XLDN2-5,a carbazole-degrading microorganism with EET ability,was selected from oilcontaminated soil as the anode-generating microorganism.The degradation and synchronous capacity of pollutants in MFC and the mechanism of EET were studied in detail.The main results are described in detail as follows:(1)Efficient treatment of breeding wastewater in microbial fuel cellBreeding wastewater served as MFC anodic chamber fuel,an electroactive microbial community that can effectively treat breeding wastewater and produce electricity was obtained by electrodomestication screening in MFC.Good power generation(power density of 438.22 mW m-2 and open-circuit voltage of 746 mV)and high pollutants removal efficiency(chemical oxygen demand of 70.05%and ammonia nitrogen of 77.43%)were obtained during the breeding wastewater treatment process in MFC.The morphology characterization by scanning electron microscope(SEM)demonstrated that the screened electroactive microbial community were able to form a thick biofilm on the anode of the MFC,and there were many kinds of microorganisms with different shapes.Microbial community analysis revealed that the microorganisms associated with electricity generation and pollutant degradation were significantly increased such as Proteiniphilum from 4.5%to 11.0%,Prolixibacteraceae from 0.06%to 8.0%,and Novosphingobium from 0.09%to 4.5%,while the pathogenic microorganisms were greatly decreased such as Paeniclostridium from 33.3%to 0.03%,Romboutsia from 29.2%to 0.01%,and Corynebacterium from 8.4%to 0.1 5%process after a round of MFC electricity-generating acclimation.(2)The carbazole degradation and simultaneous electricity generation by S.yanoikuyae XLDN2-5 in MFCS.yanoikuyae XLDN5-2 is an obligate aerobe that can grow and reproduce using nitrogen heterocyclic pollutants as carbon source,nitrogen source,and energy source.MFC was constructed with S.yanoikuyae XLDN2-5 as anodic microorganism and MSM medium containing carbazole as anolyte.From the discharge curves of the MFC during the entire operation process,it can be seen that the MFC had a short start-up period(50 h),and the continuous stable discharge time reached 700 h.The maximum discharge voltage of the MFC was 497 mV.The calculation showed that the open circuit voltage of the MFC can reach 804 mV,and the maximum power density can reach 496.8 mW m-2.The results of high-performance liquid chromatography(HPLC)showed that the concentration of carbazole in the anolyte(80 mg L-1)decreased rapidly within 24 h.and the degradation efficiency reached 64.96%within 24 h,After 24 h,the degradation rate of carbazole tended to decrease slowly,and the residue of carbazole could not be detected by HPLC after 144 h.The results of cyclic voltammetry(CV)showed that no redox reaction occurred on the surface of the anode biofilm,while electrochemical impedance spectroscopy(EIS)results showed that the formation of the anodic biofilm reduced the positive mechanism resistance.(3)Study on the EET based on redox mediators of S.yanoikuyae XLDN2-5The anolyte of S.yanoikuyae XLDN2-5 at different periods of MFC discharge and the culture medium of the entire process of shaking flask culture were measured by CV method.The results showed that there were no obvious redox peaks in the CV curves of MFC anolyte and shake flask culture medium.In addition,the experimental results of exogenous addition of redox mediators,such as thionine acetate,disodium anthraquinone-2,6-disulfonate,VB2,and 1-phenazinecarboxylic acid,showed that the output voltages of MFCs all exhibited decreases to varying degrees rather than increase.The decrease is irrelevant whether the exogenously added substances are natural substances or chemically synthesized.These above results indicated that the electron transport pathway mediated by redox mediators in MFC is not the main EET mode of S.yanoikuyae XLDN2-5.(4)Study on the EET based on conductive nanowires of S.yanoikuyae XLDN2-5SEM was used to characterize the anodic biofilms formed by S.yanoikuyae XLDN2-5 in MFC and free S.yanoikuyae XLDN2-5 cells cultured in shake flasks.SEM images showed that a large number of nanowires with an average diameter of about 50 nm were produced on the surface of S.yanoikuyae XLDN2-5 cells.Further,these nanowires could form a network structure and were cross-linked between bacterial cells in the anodic biofilm compared with free bacteria.The anodic biofilms of the MFC at different discharge periods were characterized by SEM to explore the growth process of nanowires.The results showed that a small number of nanowires have appeared on the surface of S.yanoikuyae XLDN2-5 in the anodic biofilm at the start-up discharge period of the MFC.When the discharge reached the steady period,the number of the nanowires increased dramatically and interconnected into a network.Compared with the steady discharge period,the morphologies of the anodic biofilmis at the declining discharge period and the end discharge period of the MFC showed that no significant change was observed on the number of nanowires.The growth process of nanowires was consistent with the whole discharge process of the MFC.Moreover,conductive atomic force microscope(C-AFM)further proved that the nanowires generated by S.yanoikuyae XLDN2-5 cells in the anodic biofilm have good electrical conductivity and can realize electron transf’er.(5)Study on the EET molecular mechanism of S.yanoikuyae XLDN2-5According to the above results,it is speculated that S.yanoikuyae XLDN2-5 can use EET to transfer electrons from intracellular to extracellular when growing in the low dissolved oxygen zone in the anodic biofilm to meet its own energy metabolism requirements.Transcriptome sequencing was performed for S.yanoikuyae XLDN2-5 cells cultured in shaker flask and anodic biofilms of the MFC,and target genes were screened according to gene functional annotation and gene expression.Among genes screened that may be related to EET,17 genes were knocked out according to the principle of homologous recombination,and 5 genes(SYA RS0125545,cytochrome c biogenesis protein DipZ;SYA_RS0125805,energy transducer TonB;SYA_RS0120060,flagellar protein PilZ;SYA_RS0115340,pilus assembly protein FilS;SYA_RS0102800,ubiquinol-cytochrome c chaperone)were successfully obtained knockout strains.Characteristics analysis showed that although these 5 knockout strains had lost the ability to degrade carbazole,their electricity generation performance(generation cycle,output voltage,etc.)and nanowire generation capacity in MFC were not significantly different from those of wild strains.Therefore,it is speculated that these genes were unrelated to the nanowire-based EET of S.yanoikuyae XLDN2-5 in MFC.In summary,a microbial community that can effectively treat breeding wastewater and simultaneously generate electricity in MFC was obtained in this paper.S.yanoikuyae XLDN25,an obligate aerobic carbazole-degrading bacterium,was found for the first time to be a novel electrochemically active bacterium,which can achieve pollutant degradation and simultaneous electricity generation in MFC.The EET analysis results confirmed that S.yanoikuyae XLDN25 mainly relied on microbial nanowires rather than redox mediators for its EET in MFC.The above research results can not only provide a theoretical basis for the study of EET of aerobic bacteria,but also have important significance for environmental pollution control and clean energy production using bioelectrochemical system. |
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